Fibrin glue without fibrinogen and biosealant compositions and methods

ABSTRACT

The invention is a fibrin glue that avoids the use of fibrinogen and thus eliminates the need for premixing and premature clot formation. The fibrin glue of the invention comprises thrombin, thromboplastin and calcium and may have clotting Factors, VII, IX and X, and the like. The invention also comprises a biosealant for use with the fibrin glue without fibrinogen or for use alone. The biosealant is a two component mixture of gelatin/resorcinol and glyoxal/glutaraldehyde/4-(p-maleimidophenyl) butyric acid. The two components are mixed on use.

RELATED APPLICATION

This application claims priority to application Ser. No. 60/064,864,filed Sep. 26, 1997, incorporated by reference herein.

BACKGROUND OF THE INVENTION

1. Field of the Invention

The present invention describes various bioadhesive sealants. One is ahemostatic agent commonly known as a “fibrin glue” and is a novel fibringlue that lacks fibrinogen. There is also provided a biosealantcomposition. Methods of use and manufacture of the fibrin glue andbiosealant are also provided. The fibrin glue and biosealantcompositions and methods of the present invention are suitable forarresting blood flow, maintaining hemostasis, and for accelerating andameliorating the healing process after various types of surgical andnonsurgical procedures or wound healing in mammals, including humans.

2. Description of the Related Art

The use of fibrin glue has been explored in various surgical disciplinesas reported by Lerner, R. & Binur, N. S., J. Surg. Res., 48:165-181(1990); Gibble, J. W. & Ness, P. M., Transfusion 30:741-747 (1990);Sierra, D. H., J Biometer. Applic. 7:309-352 (1993); Brennan, M., BloodReviews, 5:240-244 (1991); Dresdale, A., et al., Surgery 97:750-755(1985); Sponitz, W., et al., Amer. Surg, 59:460-462 (1987); Schlag, G. &Redl, H. (Eds), GYNECOLOGY AND OBSTETRICS-UROLOGY (1986); FIBRIN SEALANTIN OPERATIVE MEDICINE, vol 3. Springer Verlag (Berlin); and BurnouRadosevich, M. et al., Vox Sang, 58:77-84 (1990).

For example, surgeons, dentists and hematologists have reported thatfibrin glue is an effective bioadhesive. Experience in animals andhumans suggests that an advantage of using fibrin glue rather thansynthetic plastics (e.g., cyanoacrylate) or sutures is that fibrin gluepromotes local coagulation, thereby preventing bleeding even inhemophiliacs. Fibrin glue also appears to support regrowth of new tissueand the extracellular matrix.

In the past, fibrin glue has been formed by mixing two components,exogenous human fibrinogen (obtained from a source other than thepatient being treated, such as a freeze-dried plasma protein concentrateof fibrinogen/Factor XIII/fibronectin) and an activating enzyme, such asthrombin. Prior to use, the plasma protein concentrates wereconventionally solubilized in the presence of calcium chloride. Thrombininduced activation of fibrinogen resulted in the formation of fibrin.Factor XIII and calcium participated in the cross-linking andstabilization of the fibrin to produce a tight mesh of polymeric fibringlue. Applied to tissue, the fibrin clot adhered to the site ofapplication. The rate of coagulation and mechanical properties of theclot were dependent on the concentration of fibrinogen and thrombin.

Traditional fibrin glue preparations are described in InternationalApplications No. WO93/05067 to Baxter International, Inc.; WO92/13495 toFibratek, Inc.; and WO91/09641 to Cryolife, Inc.

Thrombin is a common physiological instigator of clotting. Thrombin froma number of mammalian sources, most commonly bovine, is routinely usedin commercially-available fibrin glues. Human thrombin can be employedin the formulation of fibrin glue, as can other appropriate catalyzingenzymes, such as reptilase or selected venoms (Fenton II, J. W. et al.,J Biol. Chem., 252:3587-3598 (1977); Gaffney P. J. et al., Thrombos.Haemostas., 67:424-427 (1992); European Patent Application No. EP 0 439156 A1, Stocker K., et al., Toxicon, 20:265-273 (1982); and Pirkle H. &Stocker K., Thrombos Haemostas. 65:444-450 (1991)).

Fibrinogen may be in an intimate admixture with other proteins that aretypically found in uncoagulated whole blood, in platelet-rich plasma, inplasma, in cryoprecipitate, or in precipitates of plasma obtained by amethod such as Cohn precipitations of plasma. Such additional proteincomponents may include, for example, fibronectin, immunoglobulin,particularly, IgG, and plasminogen.

Thrombin is derived from blood plasma by the fractionation of plasma.Comprehensive reviews on the preparative techniques of each have beenpublished and are the basis for most commercial plasma fractionationprocedures used by those skilled in the art (Fenton II, J. W. supra;Gaffney P. J. supra; EP 0 439 156 A1; and U.S. Pat. No. 5,143,838).

However, the prior art fibrin glue compositions required the mixture ofthrombin with exogenous fibrinogen to form an unwanted, premature fibrinclot, prior to the application of the fibrin glue to the tissue beingtreated. For example, U.S. Pat. No. 5,607,694 issued to Marx on Mar. 4,1997, discloses a fibrin glue composition in which exogenous fibrinogenand thrombin are mixed together prior to the application of the mixtureto the tissue being treated. Marx further teaches the addition ofliposomes to the fibrin glue mixture for the delivery of agents to thetissue being treated.

Similarly, U.S. Pat. No. 5,290,552 issued to Sierra et al. on Mar. 1,1994, discloses a surgical adhesive material comprising a composition offibrinogen, collagen, thrombin and calcium, in which the thrombin ismixed with exogenous fibrinogen prior to the application of the fibringlue to the tissue.

Therefore, there exists a need for a fibrin glue that can be applieddirectly to the tissue being treated to form a clot on contact withoutthe use of added exogenous fibrinogen and without pre-mixing thefibrinogen to the fibrin glue.

SUMMARY OF THE PRESENT INVENTION

The invention is a novel fibrin glue without fibrinogen that hasthromboplastin, thrombin, and calcium, together in a pharmaceuticallyacceptable carrier, such as water.

The fibrin glue without fibrinogen of the invention may comprise of0.0001 to 99.99% thromboplastin, 0.00001 to 10,000 U/ml thrombin andabout 0.015-0.025 M calcium, but preferably, there is about 0.5-1.5% orabout 1% thromboplastin, about 250-1000 U/ml thrombin, and 0.02 Mcalcium.

The fibrin glue without fibrinogen of the invention may contain otherclotting factors, growth factors, antibiotics, trace metals, etc, aslong as it contains no significant amounts of fibrinogen which wouldcause the premature formation of a clot. In a preferred embodiment, thefibrin glue also contains Factor VII, Factor IX, and Factor X. There canbe about 0.001-1000 U of each of these Factors.

In yet another embodiment, the fibrin glue without fibrinogen isformulated in a form selected from the group consisting of a bandage,surgical dressing, wound packing, swab, liquid, aerosol, paste,ointment, foam, gel, emulsion, powder and moldable form, and the like.

Methods of manufacturing the novel glue are also provided. For example,one method is: isolating prothrombin from blood, converting the isolatedprothrombin to thrombin, isolating thromboplastin from blood. Next, themethod involves blending the prothrombin and/or thrombin andthromboplastin plus calcium in a pharmaceutically acceptable carrier toproduce the fibrin glue without fibrinogen. The prothrombin may beconverted to thrombin before blending or after blending with the othercomponents of the glue. In one specific method of isolating prothrombinfrom blood described herein, the procedure also produces Factor VII,Factor IX and Factor X. Thus, the fibrin glue of the invention alsocomprises at least these additional Factors without requiring additionalpreparative techniques.

In another embodiment of the invention, the method of manufacturing afibrin glue without fibrinogen involves blending thrombin,thromboplastin and calcium in a pharmaceutically acceptable carrier toform a fibrin glue without fibrinogen. The method may also involveblending Factor VII, Factor IX and Factor X. The thrombin,thromboplastin, Factor VII, Factor IX and Factor X may be producedthrough recombinant DNA techniques or may be purified from blood orother suitable tissue source.

The invention also teaches methods of using the fibrin glue withoutfibrinogen. Because the novel glue does not pre-clot and does notrequire the mixing of components prior to or simultaneously with use, itis particularly suitable for use in surgical procedures where the gluecan be applied to an internal portion of a patient via a tube, such asan endoscope, or via a syringe, and the like. Alternatively, the gluecan be administered with devices such as a bandage, surgical dressing,wound packing, swab, syringe, tubing, endoscope, spray bottle, andaerosol canister, and the like.

The fibrin glue of the present invention is a bioadhesive that utilizesthe clotting factors of the organism being treated (the “host”),including the host's fibrinogen naturally present at a wound, to form aclot on contact with the tissue being treated to arrest blood flow.Unlike other coagulation compositions of the past, the fibrin glue ofthe present invention contains thrombin, thromboplastin and calcium andis not mixed with fibrinogen prior to application to the tissue beingtreated. The glue of the present invention also contains Factors VII, IXand X which are co-produced in the procedure employed in the manufactureof the thrombin.

In the fibrin glue of the present invention, the host system'sfibrinogen reacts with the applied thrombin/thromboplastin/calciumcomposition to form a fibrin clot at the site being treated to arrestblood flow. The fibrin glue of the present invention essentially acts asa catalyst to the host's natural clotting factors to form a fibrin cloton contact with the tissue being treated. The fibrin clot does not beginto form until the fibrin glue of the present invention is applied to thetissue being treated.

The fibrin glue without fibrinogen of the instant invention providesseveral advantages. One important advantage is that it arrests bloodflow immediately on contact with the tissue being treated and preventsunwanted, premature clot formation prior to the application to thetissue being treated. As a result, the fibrin glue may be utilized inendoscopic surgical procedures, in which the fibrin glue is appliedthrough an endoscope to access the tissue being treated, such as aninternal organ. Endoscopic use of the fibrin glues of the past provedunworkable as the premixed fibrinogen and thrombin composition wouldform a clot prior to arriving at the tissue being treated. This wouldclog the endoscope and prevent effective delivery of the fibrin glue.Also unworkable was the use of low concentrations of thrombin as thethrombin would be further diluted through the endoscope to aconcentration that was inadequate to form proper clotting of the tissuebeing treated.

The fibrin glue of the present invention may be incorporated in hydrousor anhydrous forms into bandages, dressings, and packing materials forlarge wounds, as part of first aid kits for domestic, industrial ormilitary applications, or wherever it is desired to meld biologicaltissues together or meld a biological tissue to a synthetic device. Thefibrin glue of the present invention can also be delivered through asyringe as an alternative to cauterization.

The fibrin glue may also be formulated with a pharmaceuticallyacceptable carrier in a dry delivery form such as bandages, surgicaldressings, wound packings, swabs such as Q-Tips, etc. The fibrin gluemay alternatively be formulated to be presented in liquid, aerosol, gel,emulsions, paste, ointments, foam and moldable forms.

Liquids may be delivered internally, for example, through syringes ortubing such as fiber optic tubing or endoscopes. Of course liquids maybe applied to any surface location as well. Aerosols may be deliveredfrom spray bottles, tubing and aerosol canisters. Foams may be comprisedof the fibrin glue formulated with maltodextrose, dextran or otherstarches, albumin and a surfactant. Moldable forms may be comprised ofthe fibrin glue formulated in maltodextrose, dextran or other starchesand gelatin. Gels and pastes can be formed with suitable thickeners andemulsions are well known in the art. Powders may comprise almost pureingredients or contain suitable fillers, excipients, and the like.Aerosols can be formed from liquid forms and suitable dispersants. Allof these formulation and/or delivery means are well known in the art andneed not be detailed herein.

The present invention affords a new generation of fibrin glue whoseadvantages and uses will become apparent from the following disclosureof the present invention. The present invention establish a safe andunique fibrin glue formulation for widespread use and numerous surgicaland nonsurgical applications.

Another embodiment of the invention is a biosealant that can be usedalone, or with the fibrin glue of the invention. The biosealant has afirst component with gelatin and resorcinol in water and a secondcomponent with glyoxal, 4-(p-maleimidophenyl) butyric acid, andglutaraldehyde in water. The first and second components are admixedjust prior to, or simultaneously with, application to the patient.

More particularly, the first component has 0.06-10 g gelatin per 500 mland 0.001-5 M resorcinol, and the second component has 0.001-10 Mglyoxal, 1-99% glutaraldehyde and 0.001 mg-10 g/100 ml of4-(p-maleimidophenyl) butyric acid. However, in a preferred embodimentthe first component has about 0.75 g/liter gelatin and about 0.125 Mresorcinol, the second component has about 33.3% glyoxal, about 5%glutaraldehyde and about 50 mg/liter 4-(p-maleimidophenyl) butyric acid.Varying the ratio of the first and second components determines the timerequired for polymerization. Thus, the components may be mixed in anydesired ratio, including for example, a 1:0.01 to a 1:0.15 ratio or a1:0.20 ratio (component 1:component 2).

The improved biosealant of the invention has a greater tensile strengththan fibrinogen alone. It can be used in heart valves, skin grafts, forparenchymal tissue such as liver, spleen, lung, spinal cord and othernervous system tissues, cosmetic surgery, as an orthopedic adhesive, orfor any other application where it is desired to meld biological tissuestogether or to meld biological tissue to a synthetic device.

DESCRIPTION OF THE PRESENT INVENTION

The invention can be best understood by reference to the followingdefinitions:

Biosealant

A sealant that is appropriate for biological use wherever it is desiredto meld biological tissues (or a tissue and an implant) together. Thebiosealant of the present invention is a two-component system that uponadmixing polymerizes to form a seal with high tensile strength. Thefirst component has at least gelatin, resorcinol or their equivalents inwater. The second component has at least glyoxal and glutaraldehyde and4-(p-maleimidophenyl) butyric acid or their equivalents in water.

Fibrin Glue

A term of art that refers to a bioadhesive employing the fibrin clottingcomponents.

Fibrin Glue without Fibrinogen

The bioadhesive of the invention which lacks significant amounts offibrinogen, and instead employs the in situ fibrinogen found in the hostto begin the clotting cascade. The fibrin glue without fibrinogen has atleast thrombin, thromboplastin and calcium, and preferable also containsclotting Factors VII, IX and X.

Pharmaceutically Acceptable Carrier

Any carrier, including water, that can be used to formulate acomposition for medicinal uses. This term is well known in the art andneed not be described further herein.

TAPSO

3-[N-tris (hydroxymethyl) methylamino]-2-hydroxy-propanesulfonic acid.

Thrombin

Any human, mammalian or synthetically produced thrombin or itsequivalent, such as snake venom catalyzing enzymes, thrombin precursorsand any derivatives or muteins that are biologically active. Thrombinproduced from natural sources may contain other components therein,including other clotting pathway components (except fibrinogen in anysignificant amount), especially clotting Factors VII, IX and X.

Thromboplastin

Any human, mammalian or synthetically produced thromboplastin or itsequivalent, such as precursors, and derivatives or muteins that arebiologically active.

The fibrin glue of the present invention comprises a composition ofthrombin and thromboplastin. Factors VII, IX, X and calcium may also beincluded. Set forth below is a description of steps for preparing oneembodiment of the fibrin glue without fibrinogen of the presentinvention. Suitable ranges for elements are provided in parenthesis.

EXAMPLE 1 Manufacture of Thrombin, Factors VII, IX and X

Thrombin and Factors VII, IX, and X are co-produced in the followingprocedure:

1. To human 3.8% sodium citrate plasma, add slowly 10% by volume 1 MBaCl₂ (range 0.01-2.0 M, preferred range 0.8-1.2 M), while mixing gentlyat 2-8° C.

2. Mix for two hours.

3. Centrifuge at 5,000 rpm for ten minutes at 2-8° C.

4. Discard supernatant and retain precipitate.

5. Resuspend precipitate with 10% of the starting plasma using 40%(NH₄)₂SO₄ (range 1-65%, preferred range 30-50%), and gently mix for16-24 hours.

6. Centrifuge at 5,000 rpm for ten minutes at 2-8° C.

7. Discard precipitate and retain supernatant.

8. To the super add slowly 22 g (NH₄)₂SO₄/100 ml (range 1-35 g/100 ml,preferred range 18-26 g/100 ml) supernatant while mixing.

9. Mix for 2 hours at 2-8° C.

10. Centrifuge at 12,000 rpm for 3 hours at 2-4° C.

11. Discard supernatant and retain precipitate.

12. Resuspend precipitate with dialysis buffer (0.5 M TAPSO, pH7.6±0.02, 0.9% NaCl) (range 0.01-1 M TAPSO, pH 4-10, 0.5-1.5% NaCl,preferred range 0.4-0.6 M TAPSO, pH 7-8, 0.8-1.1% NaCl) in about 10% byvolume from step 8.

13. Dialyze in a 1000 MW cellulose acetate tubing for 18-24 hours,changing the dialysis buffer two times.

14. Convert the isolated prothrombin to thrombin by adding 0.4-0.6 gsodium citrate per ml (suitable range 0.01-1.0 g/ml, preferred range0.3-0.9 g/ml) and seeding it with 1000 U thrombin per ml of prothrombin(suitable range 1-10,000 U/ml, preferred range 250-2000 U/ml).

15. Incubate for 24 hours at 37° C.

EXAMPLE 2 Manufacture of Thromboplastin

Thromboplastin is isolated by the following procedure:

1. Concentrate platelets by filtering off the plasma through a Buchnerfunnel.

2. Wash platelets with 0.9% NaCl (suitable range 0.1%-2%).

3. Resuspend platelets with cold 0.9% NaCl in two times volume.

4. Place in a blender for ten minutes at high speed (avoid temperaturerising above 37° C.).

5. Centrifuge at 5,000 rpm for ten minutes at 2-8° C.

6. Discard precipitate and retain supernatant.

EXAMPLE 3 Blending the Fibrin Glue

In order to manufacture the fibrin glue of the invention the followingcomponents are admixed.

1. Add 1% by volume (range 0.0001-99.99%, preferred range 0.5-1.5%) ofthe thromboplastin from Example 2, step 6 to the thrombin concentratefrom Example 1, step 15 (range 0.00001-100,000 U/ml, preferred range1-100,000 U/ml).

2. Add CaCl₂ to the thrombin/thromboplastin solution to a concentrationof 0.02 M (range 0.0-1M, preferred range 0.015-0.025 M).

3. Blend with a pharmaceutically acceptable carrier if desired. Suchcarriers and blending techniques are well known in the art and need notbe described herein.

It is recognized that prothrombin, thromboplastin, and Factors VII, IXand X, for use in the composition of the fibrin glue of the presentinvention can be obtained from other than a human source, such as fromanimals, or may be synthetically produced, such as with recombinant DNAtechniques known by those skilled in the art. Further, because of thegeneral risks and problems of infections from pooled blood products, thehuman blood plasma used as a source for the prothrombin andthromboplastin should be tested for contaminates such as lipid-envelopedviruses such as HIV and HCV (also known as non A-non B hepatitis virus),as well as cytomegalovirus (CMV), Epstein-Barr virus, and the herpessimplex viruses.

EXAMPLE 4 Manufacture of Sealant

Another embodiment of the fibrin glue of the present invention comprisesan improved sealant or “biosealant” with a greater tensile strength thanfibrinogen used alone as a sealant. The improved biosealant of thepresent invention is a two-component system that provides a 200 g/cm²tensile strength in the first minute after application to the site beingtreated, and 650 g/cm² two hours after application. This is a greatadvantage over a tensile strength of 50 g/cm² and 450 g/cm²,respectively, using fibrinogen.

The first component of the improved sealant comprises 375 ml of 1 ggelatin/500 ml H₂O, 125 ml of 1 M resorcinol, and 500 ml of H₂O (range0.06-10 g gelatin per 500 ml and 0.001-5 M resorcinol, preferred rangeabout 0.2-0.5 g gelatin/500 ml and about 0.05-2 M resorcinol).

The second component of the improved sealant (for a volume of 250 ml forexample), comprises 225 ml of 37% (1.76 M) glyoxal and 25 ml of 50%aqueous solution of glutaraldehyde containing 50 mg/100 ml of4-(p-maleimidophenyl) butyric acid (range 0.001-10 M glyoxal, 1-99%glutaraldehyde and 0.001 mg-10 g/100 ml of 4-(p-maleimidophenyl) butyricacid, preferred range about 1-2 M glyoxal, about 40-60% glutaraldehydeand about 40-60 mg/100 ml of 4-(p-maleimidophenyl) butyric acid).

The improved sealant is formed from the polymerization of 0.15 ml of thesecond component to each 1 ml of the first component, which takesapproximately 1.5 minutes for full reaction after the first and secondcomponents are thoroughly mixed. By varying the ratio of the twocomponents it is possible to effectively control the reaction time forpolymerization without losing tensile strength. For example, a 45 secondsealant is produced with a 0.2:1 ratio of second component to first. A3-4 minute sealant is obtained with a 0.05:1 ratio.

While the present invention has been described in detail with respect toits preferred embodiment, it is appreciated that other variations of thepresent invention may be devised which do not depart from the inventiveconcept of the present invention. All art cited herein is expresslyincorporated by reference and is re-listed here for convenience only.

Brennan, M., Blood Reviews, 5:240-244 (1991). Burnou Radosevich, M. etal., Vox Sang, 58:77-84 (1990). Dresdale, A., et al., Surgery 97:750-755(1985). European Patent Application No. EP 0 439 156 A1. Fenton II, J.W. et al.; j Biol. Chem., 252:3587-3598 (1977). Fibrin Sealant inOperative Medicine, vol 3. Springer Verlag (Berlin). Gaffney P. J. etal., Thrombos. Haemostas., 67:424-427 (1992). Gibble, J. W. & Ness, P.M., Transfusion 30:741-747 (1990). Lerner, R. & Binur, N. S., J. SurgRes., 48:165-181(1990). Pirkle H. & Stocker K., Thrombos. Haemostas.65:444-450 (1991). Schlag, G. & Redl, H. (Eds), GYNECOLOGY ANDOBSTETRICS- UROLOGY (1986). SIERRA, D. H., J BIOMETER. APPLIC. 7:309-352(1993). SPONITZ, W., ET AL., AMER. SURG., 59:460-462 (1987). STOCKER K.,ET AL., TOXICON, 20:265-273 (1982). U.S. PAT. NO. 5,143,838. U.S. PAT.NO. 5,607,694. U.S. PAT. NO. 5,290,552. U.S. PAT. NO. 5,185,001 U.S.PAT. NO. 5,219,328 U.S. PAT. NO. 5,385,606 U.S. PAT. NO. 5,405,607 U.S.PAT. NO. 5,589,462 U.S. PAT. NO. 5,605,541 U.S. PAT. NO. 5,607,694 U.S.PAT. NO. 5,610,147 WO93/05067. WO 92/13495. WO 91/09641.

What is claimed is:
 1. A fibrin glue without fibrinogen, comprising: a)thromboplastin, b) thrombin, and c) calcium, together in apharmaceutically acceptable carrier.
 2. The fibrin glue withoutfibrinogen of claim 1, wherein there is from 0.0001 to 99.99%thromboplastin, 0.00001 to 10,000 U/ml thrombin and about 0.02 Mcalcium.
 3. The fibrin glue without fibrinogen of claim 1, furthercomprising Factor VII, Factor IX, and Factor X.
 4. The fibrin gluewithout fibrinogen of claim 2, further comprising Factor VII, Factor IX,and Factor X.
 5. The fibrin glue without fibrinogen of claim 3, whereinthere is from about 0.001-1000 U/ml Factor VII, about 0.001-1000 U/mlFactor IX, and about 0.001-1000 U/ml Factor X.
 6. The fibrin gluewithout fibrinogen of claim 4, wherein there is from about 0.001-1000U/ml Factor VII, about 0.001-1000 U/ml Factor IX, and about 0.001-1000U/ml Factor X.
 7. The fibrin glue without fibrinogen of claim 1, whereinthe fibrin glue without fibrinogen is formulated in a form selected fromthe group consisting of a bandage, surgical dressing, wound packing,swab, liquid, aerosol, paste, ointment, foam, gel, emulsion, powder andmoldable form.
 8. The fibrin glue without fibrinogen of claim 3, whereinthe fibrin glue without fibrinogen is formulated in a form selected fromthe group consisting of a bandage, surgical dressing, wound packing,swab, liquid, aerosol, paste, ointment, foam, gel, emulsion, powder andmoldable form.
 9. A fibrin glue without fibrinogen, consistingessentially of: a) thromboplastin, b) thrombin, and c) calcium, togetherin a pharmaceutically acceptable carrier.
 10. The fibrin glue withoutfibrinogen of claim 9, wherein there is from about 0.0001 to 99.99%thromboplastin, about 0.00001 to 10,000 U/ml thrombin and about0.015-0.025 M calcium.
 11. A fibrin glue without fibrinogen, consistingessentially of: a) thromboplastin, b) thrombin, and c) calcium, d)Factor VII, e) Factor IX, and f) Factor X.
 12. The fibrin glue withoutfibrinogen of claim 11, wherein there is from about 0.0001 to 99.99%thromboplastin, about 0.00001 to 10,000 U/ml thrombin, about 0.015-0.025M calcium, about 0.001-1000 U/ml Factor VII, about 0.001-1000 U/mlFactor IX, and about 0.001-1000 U/ml Factor X.
 13. The fibrin gluewithout fibrinogen of claim 11, wherein there is from about 0.5-1.5%thromboplastin, about 250-1000 U/ml thrombin, about 0.02 M calcium,about 0.001-1000 U/ml Factor VII, about 0.001-1000 U/ml Factor IX, andabout 0.001-1000 U/ml Factor X.
 14. The fibrin glue without fibrinogenof claim 11, wherein the fibrin glue without fibrinogen is formulated ina form selected from the group consisting of a bandage, surgicaldressing, wound packing, swab, liquid, aerosol, paste, ointment, foam,gel, emulsion, powder and moldable form.
 15. A method of using a fibringlue without fibrinogen, comprising administering said fibrin gluewithout fibrinogen to an internal portion of a patient by a tube orsyringe.
 16. A method of using a fibrin glue without fibrinogen,comprising administering said fibrin glue without fibrinogen to apatient with a device selected from the group consisting of a bandage,surgical dressing, wound packing, swab, syringe, tubing, endoscope,fiber optic tubing, spray bottle, foam moldable form and aerosolcanister.